Whole blood is made up of various cellular components such as red cells, white cells and platelets suspended in its liquid component, plasma. Whole blood can be separated into its constituent components (cellular or liquid), and the desired separated component can be administered to a patient in need of that particular component. For example, platelets can be removed from the whole blood of a healthy donor, collected, and later administered to a cancer patient whose ability to “make” platelets has been compromised by chemotherapy or radiation treatment.
Commonly, platelets are collected by introducing whole blood into a centrifuge chamber wherein the whole blood is separated into its constituent components, including platelets, based on the size and densities of the different components. This requires that the whole blood be passed through a centrifuge after it is withdrawn from, and before it is returned to, the donor. Typical blood processing systems thus include a permanent, reusable centrifuge assembly containing the hardware (drive system, pumps, valve actuators, programmable controller, and the like) that spins and pumps the blood, and a disposable, sealed and sterile fluid processing assembly that is mounted cooperatively on the hardware. The centrifuge assembly spins a disposable centrifuge chamber in the fluid processing assembly during a collection procedure, thereby separating the blood into its constituent components.
Automated blood separation systems are used to collect large numbers of platelets. Automated systems perform the separation steps necessary to separate platelets from whole blood in a sequential process with the donor present. Automated systems draw whole blood from the donor, separate out the desired platelets from the drawn blood, and return the remaining red blood cells and plasma to the donor, all in a sequential flow loop. Large volumes of whole blood can be processed using an automated on-line system. Due to the large processing volumes, large yields of concentrated platelets can be collected. Moreover, since the donor's red blood cells are returned, the donor can donate platelets for on-line processing much more frequently.
In the automated, on-line separation and collection of platelets, sometimes referred to as platelet apheresis, or simply “plateletpheresis”, the platelets are separated from whole blood and concentrated in the centrifuge chamber or elsewhere in the fluid processing set (hereinafter “platelet concentrate” or “PC”). Although most of the plasma is removed during apheresis, a volume of plasma still remains in the PC, hereinafter referred to as “residual plasma”. (Platelets may also be derived from buffy coats obtained from manually collected units of whole blood. A plurality of buffy coats are typically pooled to provide an amount or dose of platelets suitable for transfusion.) The platelets, whether collected by apheresis or from pooled buffy coats, are typically reconstituted in a liquid storage medium, such as plasma and/or a synthetic storage solution, for storage until needed for transfusion to a patient.
For the stored platelets to be suitable for later administration, they must substantially retain their viability and platelet function. A number of interrelated factors may affect platelet viability and function during storage. Some of these factors include the anticoagulant used for blood collection, the method used to prepare the platelets, the type of storage container used, and the medium in which the platelets are stored.
Currently, platelets may be stored for five or even seven days at 22° C. After seven days, however, platelet function may become impaired. In addition to storage time, other storage conditions have been shown to affect platelet metabolism and function including pH, storage temperature, total platelet count, plasma volume, agitation during storage and platelet concentration.
A variety of assays have been developed which attempt to determine the quality of stored platelets and the in vivo viability of those platelets when transfused to a patient. For instance, the percentage of platelets that respond appropriately to an ADP agonist (the ESC assay) and the percentage of platelets that respond appropriately to hypotonic shock (HSR assay) are two assays which are thought to correlate well with viability of stored platelets. ESC is a photometric assessment of discoid platelet shape change in response to an ADP agonist. VandenBroeke, et al., “Platelet storage solution effects on the accuracy of laboratory tests for platelet function: a multi-laboratory study.” Vox Sanguinis (2004) 86, 183-188.
The results of the HSR (Hypotonic Shock Response) assay are often considered to correlate strongly with the in vivo effectiveness of platelets when they are introduced into an individual. This assay measures the ability of platelets to recover a discoid shape after swelling in response to a hypotonic environment. Higher scores on either the HSR or ESC assay appear to correlate with increased viability of the platelets when transfused to patients. The methods and uses of the HSR and ESC assays are described in more detail by Holme et al. Transfusion, January 1998; 38:31-40, which is incorporated by reference herein.
Another assay for measuring platelet viability based on platelet shape is the Morphology Score for the platelets during and after storage. Morphology Scores may be determined by, for example, the Kunicki Method, whereby a selected number of platelets in a sample are examined to determine the shape, e.g., discoid, spherical, dendritic or balloon. The number of each shape is then multiplied by a selected multiplier and the resultant numbers are summed to arrive at a Morphology Score. A score of 250-400 is typically indicative of a viable platelet population.
The presence of the glycoprotein P-selectin on the surface of platelets is also used to characterize the viability of platelets upon transfusion with the presence of P-selectin believed to indicate a loss of viability. As described by Holme et al. Transfusion 1997; 37:12-17 and incorporated herein by reference, platelets undergo a shape change transforming from disc shaped to sphere shaped upon platelet activation. This activation is thought to involve the secretion of β-thromboglobulin from the alpha granules resulting in the appearance of P-selectin on the surface of the platelets. Antibodies directed against P-selectin, such as the monoclonal antibody CD62P, are used to detect the presence of P-selectin on the surface of platelets and have been used as a marker of platelet activation and a decreased viability of the platelets upon transfusion.
A further indicator of platelet viability is pH. Stored platelets will typically produce some lactic acid (as a by-product of anaerobic glycolysis.) The increase in lactic acid gradually acidifies the storage media. This acidification of the media alters platelet physiology and morphology such that when the pH of the media drops below about 6.2 the platelets may be considered nonviable.
Although residual plasma is effective for storage of platelets, it may not be the ideal medium for platelet storage because plasma itself is a valuable blood component that can be removed from the platelets and then used or further processed for use in the treatment of patients with other disorders. Another reason for minimizing the volume of plasma from platelet concentrate is to prevent allergic transfusion reactions (ATR) to plasma. There may be other reasons for removing at least some or even most of the plasma from the platelets. For example, the presence of certain antibodies in plasma has been correlated with the occurrence of TRALI (transfusion-related acute lung injury) in some patients. Consequently, while residual plasma may be present to some degree in platelets obtained in an apheresis procedure, it may be desirable to minimize the volume of plasma and combine the residual plasma with a synthetic additive solution.
Thus, platelets may be resuspended and stored in a combined storage medium that includes a relatively small volume of residual plasma and a volume of the synthetic additive solution (AS). In accordance with certain existing protocols, the combination of plasma and a synthetic AS is provided in a pre-selected ratio of plasma to AS.
One such ratio is 35% plasma:65% AS. More recently, as described in U.S. Patent Publication Nos. US 2009/0191537 and International Patent Application Publication WO 2012/139017, the contents of which is incorporated herein by reference in its entirety, the relative percentage of plasma may be reduced further to below 35%, below 20%, and as low as approximately 10%, resulting in a combined storage medium with a plasma to AS ratio of 20:80 and 10:90, respectively.
The desired ratios are often pre-programmed into the collection apparatus (i.e., apheresis device), and the appropriate volumes of plasma and AS are combined, adjusted and sometimes re-adjusted to arrive at the pre-selected ratio. Where a high yield of platelets is collected, a greater volume of the combined storage medium may be required. This may result in including more than the desired volume of plasma in the combined storage medium.
Thus, it would be desirable to provide a storage environment for the extended storage of platelets that includes a minimal amount of plasma that is not tied to a pre-determined ratio of plasma to AS but rather utilizes a fixed volume of residual plasma that does not require adjusting.